Post-transcriptional gene silencing (PTGS), which operates at the level of sequence-specific RNA degradation, has emerged as a major phenomenon through which transgene expression in plants is down-regulated. It was first recognized in plants, and similar mechanisms since then have been observed in non-plant systems, where it is known by different names, to with, quelling in the fungus Neurospora crassa (Romano and Macino (1992) Mol Microbiol 6:3343-3353), and RNA interference (RNAi) in worms, flies, and mammals (Bosher and Labouesse (2000) Nat Cell Biol 2:E31-36).
Although the mechanism remains to be fully elucidated, it appears that double-stranded RNA (dsRNA) serve as key intermediates in PTGS (Bass (2000) Cell 101:235-238). The involvement of dsRNA is supported by identification of small complementary RNA (cRNA), 21-25 nucleotides long, which can bind the target RNA to form dsRNA, in PTG-silenced plants (Hamilton and Baulcombe (1999) Science 286:950-952), and the finding that a protein similar to RNA-dependent RNA polymerase, the enzyme involved in cRNA synthesis, is required for PTGS (Mourrain et al. (2000) Cell 101:533-542).
Another protein identified to be required for PTGS is the ARGONAUTE (AGO1) protein (Bohmert et al. (1998) EMBO J. 17:170-180; Fagard et al. (2000) Proc Nat Acad Sci USA 97:11650-11654). AGO1 protein shares homology with the RDE1 and QDE-2 proteins which have been found to be required for RNAi in C. elegans and for quelling in Neurospora, respectively, thus reinforcing the notion that PTGS, RNAi, and quelling are similar processes at the mechanistic level. AGO1/RDE1/QDE-2 proteins are similar to eIF2C, a protein important for protein translation. It is therefore hypothesized that dsRNA mediates PTGS by disrupting proper positioning of eIF2C in the translation machinery complex, thereby preventing translation of the target mRNA (Tabara et al. (1999) Cell 99:123-132; Fagard et al. (2000) Proc Natl Acad Sci USA 97:11650-11654).
It is apparent that PTGS is an important process, which if manipulated properly, may be used to control transgene expression. Disclosed herein are sequences very homologous to the AGO1 protein family, which includes the ZWILLE (ZLL) or PINHEAD (PNH) protein involved in plant development (Moussian et al. (1998) EMBO J. 17:1799-1809; Lynn et al. (1999) Development 126:469-481), and the RDE-1 protein involved in transposon silencing (Tabara et al. (1999) Cell 99:123-132). These sequences may be used to manipulate PTGS. Since some of the AGO1 family members have also been shown to be involved in transposon silencing, meristem development, and differentiation of meristematic tissue, the polynucleotides disclosed herein may also be used to manipulate transposon activity, meristem activity, plant architecture and development, and proliferation of undifferentiated plant cells in culture, which would be useful in callus propagation.